Parathyroid hormone-related protein (PTHrP) is responsible for humoral hypercalcemia of malignancy (HHM). PTHrP is produced by most tissues of the body and acts as a regulator of: a) transepithelial calcium transport; b) cellular growth, differentiation, apoptosis and organogenesis; and, c) smooth muscle tone. In vascular smooth muscle (VSM), PTHrP serves as a vasodilator, acting through the nitric oxide and adenylyl cyclase pathways. PTHrP appears to participate in the proliferative response to arterial injury, as occurs in arterial bypass surgery and coronary angioplasty. Recently, we have demonstrated that PTHrP may have directionally opposite effects on the VSM cell proliferation: when it is added exogenously to cultured VSM, it acts via the PTH/PTHrP receptor to inhibit proliferation, probably via cAMP. However, when it is introduced within the cell by gene transfer, it dramatically stimulates proliferation. This stimulation of proliferation in VSM is dependent on the presence of nuclear localization sequences within the PTHrP molecule, and is associated with the entry of PTHrP into the nuclear compartment. The studies described in the current application are designed to explore the cellular mechanisms responsible for this nuclear targeted proliferative effect. The studies use VSM as a model, but it is anticipated that the results will apply to the broad array of tissues which express PTHrP. Specifically, we plan: 1. To fully define the structural features of the PTHrP molecule required for nuclear targeting and activation of VSM proliferation. 2. To define the mRNA species and proteins which are regulated by PTHrP in the proliferative response to nuclear expression of PTHrP. 3. To explore the role of PTHrP in normal vascular development in the "rescued PTHrP knockout" mouse, and in the PTH/PTHrP receptor "knockout" mouse. These studies should help clarify the mechanism of action of PTHrP in the skeleton, the vascular tree, and the many other tissues which produce it. Further, they will lay the groundwork for the possible future use of PTHrP as a therapeutic agent for disorders of the skeleton, the cardiovascular system, and other tissues.